Arthropod-borne viruses (arboviruses) have gained in public health and veterinary importance worldwide, as demonstrated by the expansion of geographic range and rising incidence of viruses such as West Nile, Rift Valley, dengue, Chikungunya, Venezuelan equine encephalitis and Japanese encephalitis during the past decades. Drivers such as urbanization, migration, and deforestation have been implicated as underlying causes for arboviral disease emergence. Host susceptibility has also been speculated to play a key role in determining the geographic distribution of some arboviral diseases, but the contribution and mechanism of this important factor has not been well studied for arboviral diseases. It is our objective to measure the degree of cross-reactivity in human alphaviral infections which may impact host susceptibility and to elucidate the mechanisms underlying this. The central hypothesis to be tested in this project is that heterologous immunity is protective in alphaviral infection, mediated by non-neutralizing antibodies acting via Fc receptor-mediated effector functions. These mechanisms act as a bridge between the adaptive humoral response and the innate immune response, and as such, have the potential to become activated quickly to prevent disease acquisition. We propose studying these mechanisms by examining serum antibody and memory B-cell cross-reactivity to heterologous virus. We have a unique opportunity to study these mechanisms using banked specimen from Darien, Panama. In 2010, eastern equine encephalitis virus (EEEV) newly emerged in this region known to be endemic for Venezuelan equine encephalitis virus (VEEV) (both are members of the Alphavirus genus, family Togaviridae). Additional alphaviruses, Mayaro (MAYV) and the now-pandemic Chikungunya (CHIKV), have not yet emerged in this region, but are poised to do so, adding urgency to the need to understand heterologous immunity in arboviral disease. The hypothesis will be tested in two Specific Aims: 1. Determine the extent of heterologous immunity in VEEV-exposed versus alphavirus-nave subjects via FcR-mediated effector functions and neutralization; 2. Assess durability of humoral immunity by detection of antigen-specific memory B-cells (MBCs) and cross-reactive serum- and MBC-derived antibodies in longitudinal samples. This project will provide intensive career development experience in viral pathogen-host immunology with guidance from a world-class mentoring team with complementary expertise. The multi-faceted training plan includes a rigorous laboratory component, courses of international repute in immunology and infectious disease modeling, and career guidance. If successful, this study will provide novel insights into alphaviral immune responses that can be harnessed in disease modeling efforts to examine implications of heterologous immunity for disease emergence. These insights will also have direct application for monoclonal antibody therapeutics and vaccine development.